With the x-ray analysis of the structures of the serine proteases chymotrypsin A alpha and A gamma, subtilisin and elastase at high resolution, it is now possible to carry out detailed studies of the active site regions of these enzymes. We intend to prepare crystallin small molecule-enzyme complexes from these proteases and study them crystallographically. Major objectives will include preparation of a virtual acyl enzyme and a virtual substrate-enzyme complex from one or more of the above enzymes. Acyl enzymes and enzyme-substrate complexes obtained from these enzymes are not stable long enough for crystallographic measurements. We have, therefore, designed and plan to synthesize small molecules (virtual substrates) which are structurally similar to natural substrates and yet should form stable useful derivatives. Difference Fourier electron density maps will be obtained from those derivatives which prove to be isomorphous with the native proteins. Information thus should be learned about binding modes, recognition sites and the mechanism of enzymatic catalysis. Inhibitors for the acid protease from Rhizopus chinesis will be designed and synthesized for the purposes of preparing a heavy atom derivative to aid the crystallographic structure determination and to map the substrate binding site of this protease.